Role of formation of an ERK-FAK-paxillin complex in migration of human corneal epithelial cells during wound closure in vitro.

PURPOSE Migration of corneal epithelial cells is an important step in the corneal wound healing. The role of extracellular signal regulated kinase (ERK) for the regulation of cell migration during wound closure was examined. METHODS Scratch wounds were introduced into human corneal epithelial cells in the absence or presence of PD98059, an ERK signaling inhibitor. The phosphorylation and localization of ERK during wound closure were examined by immunoblot and immunofluorescence analyses. The tyrosine phosphorylation of focal adhesion kinase (FAK) and paxillin, as well as the association of FAK with paxillin and ERK, were evaluated by immunoprecipitation and immunoblot analysis. The effect of a mutant form of MEK1 on cell migration and proliferation was determined by transfection. RESULTS PD98059 inhibited cell migration in a concentration- and time-dependent manner. Wounding increased the phosphorylation of ERK as well as the tyrosine phosphorylation of FAK and paxillin in a manner sensitive to PD98059. Furthermore, wounding induced the formation of an ERK-FAK-paxillin complex and this effect as well as the wounding-induced formation of focal adhesions, membrane ruffles, and bundles of F-actin, were inhibited by PD98059. Phosphorylated ERK localized at the wound margin, and such localization was not observed in the presence of PD98059. Expression of dominant negative mutant form of MEK1 inhibited cell migration during wound closure without the effect of cell proliferation. CONCLUSIONS ERK regulates cell migration during wound healing in vitro by modulating the phosphorylation of FAK and paxillin and the consequent formation of focal adhesions. An ERK-FAK-paxillin signaling pathway may play an important role in corneal epithelial wound healing in vivo.

[1]  Giampaolo Manao,et al.  Proliferation Versus Migration in Platelet-derived Growth Factor Signaling , 2008, Journal of Biological Chemistry.

[2]  J. Capó-Aponte,et al.  Phosphatase-mediated crosstalk control of ERK and p38 MAPK signaling in corneal epithelial cells. , 2006, Investigative ophthalmology & visual science.

[3]  T. Nishida,et al.  Role of Rac1 in fibronectin-induced adhesion and motility of human corneal epithelial cells. , 2006, Investigative ophthalmology & visual science.

[4]  Satoko Nishimoto,et al.  MAPK signalling: ERK5 versus ERK1/2 , 2006, EMBO reports.

[5]  R. Baffa,et al.  Proepithelin promotes migration and invasion of 5637 bladder cancer cells through the activation of ERK1/2 and the formation of a paxillin/FAK/ERK complex. , 2006, Cancer research.

[6]  Y. Kariya,et al.  Insulin-like growth factor-1 induces migration and expression of laminin-5 in cultured human corneal epithelial cells. , 2006, Investigative ophthalmology & visual science.

[7]  R. Mohan,et al.  Wound Healing in the Cornea: A Review of Refractive Surgery Complications and New Prospects for Therapy , 2005, Cornea.

[8]  M. Karin,et al.  From JNK to Pay Dirt: Jun Kinases, their Biochemistry, Physiology and Clinical Importance , 2005, IUBMB life.

[9]  T. Chikama,et al.  Participation of p38 MAP Kinase, but Not p44/42 MAP Kinase, in Stimulation of Corneal Epithelial Migration by Substance P and IGF-1 , 2005, Current eye research.

[10]  L. Cantley,et al.  Paxillin serves as an ERK-regulated scaffold for coordinating FAK and Rac activation in epithelial morphogenesis. , 2004, Molecular cell.

[11]  Miki Ebisuya,et al.  ERK Activation Propagates in Epithelial Cell Sheets and Regulates Their Migration during Wound Healing , 2004, Current Biology.

[12]  K. Jacobson,et al.  A Role for JNK-Paxillin Signaling in Cell Migration , 2004, Cell cycle.

[13]  G. Sharma,et al.  p38 and ERK1/2 Coordinate Cellular Migration and Proliferation in Epithelial Wound Healing , 2003, Journal of Biological Chemistry.

[14]  Michael D Schaller,et al.  Paxillin: a focal adhesion-associated adaptor protein , 2001, Oncogene.

[15]  S. Ebner,et al.  Glial cell-derived neurotrophic factor (GDNF)-induced migration and signal transduction in corneal epithelial cells. , 2001, Investigative ophthalmology & visual science.

[16]  Renato Ambrósio,et al.  The Corneal Wound Healing Response: Cytokine-mediated Interaction of the Epithelium, Stroma, and Inflammatory Cells , 2001, Progress in Retinal and Eye Research.

[17]  K. Hirata,et al.  Lysophosphatidylcholine inhibits endothelial cell migration and proliferation via inhibition of the extracellular signal-regulated kinase pathway. , 2000, Arteriosclerosis, thrombosis, and vascular biology.

[18]  K. Sobue,et al.  Changes in the Balance of Phosphoinositide 3-Kinase/Protein Kinase B (Akt) and the Mitogen-activated Protein Kinases (ERK/p38MAPK) Determine a Phenotype of Visceral and Vascular Smooth Muscle Cells , 1999, The Journal of cell biology.

[19]  D. Schlaepfer,et al.  Signaling through focal adhesion kinase. , 1999, Progress in biophysics and molecular biology.

[20]  C. Nobes,et al.  Rho, Rac, and Cdc42 GTPases regulate the assembly of multimolecular focal complexes associated with actin stress fibers, lamellipodia, and filopodia , 1995, Cell.

[21]  L. Lim,et al.  The Ras-related protein Cdc42Hs and bradykinin promote formation of peripheral actin microspikes and filopodia in Swiss 3T3 fibroblasts , 1995, Molecular and cellular biology.

[22]  H. Handa,et al.  An SV40-immortalized human corneal epithelial cell line and its characterization. , 1995, Investigative ophthalmology & visual science.

[23]  Anne J. Ridley,et al.  The small GTP-binding protein rac regulates growth factor-induced membrane ruffling , 1992, Cell.

[24]  T. Nishida,et al.  Coordinated appearance of β1 integrins and fibronectin during corneal wound healing , 1992 .

[25]  J. Lanigan,et al.  Fibronectin in healing rabbit corneal wounds. , 1981, Laboratory investigation; a journal of technical methods and pathology.

[26]  D. Hammer,et al.  Integrin-mediated signalling through the MAP-kinase pathway. , 2008, IET systems biology.

[27]  J. Ramos.  The regulation of extracellular signal-regulated kinase (ERK) in mammalian cells. , 2008, The international journal of biochemistry & cell biology.

[28]  S. Saika,et al.  Role of p38 MAP kinase in regulation of cell migration and proliferation in healing corneal epithelium. , 2004, Investigative ophthalmology & visual science.

[29]  今安 正樹.  Phosphorylation of MAP kinase in corneal epithelial cells during wound healing , 2003 .

[30]  T. S. Panetti,et al.  Tyrosine phosphorylation of paxillin, FAK, and p130CAS: effects on cell spreading and migration. , 2002, Frontiers in bioscience : a journal and virtual library.

[31]  T. Awata,et al.  Fibronectin in corneal wound healing: appearance in cultured rabbit cornea. , 1982, Japanese journal of ophthalmology.

[32]  • Epidermis,et al.  WOUND healing. , 1959, The Medical journal of Australia.